scholarly journals A new organotypic model of synaptic competition reveals activity-dependent localization of C1q

2017 ◽  
Author(s):  
Ryuta Koyama ◽  
Yuwen Wu ◽  
Allison R. Bialas ◽  
Andrew Thompson ◽  
Christina A. Welsh ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Neural Circuits ◽  
Ex Vivo ◽  
Molecular Pathways ◽  
Synapse Elimination ◽  
Synaptic Competition ◽  
Coculture Model ◽  
First Time ◽  
Activity Dependent

AbstractImmature neural circuits undergo synaptic refinement, in which activity-dependent competition between synapses results in pruning of inappropriate connections and maintenance of appropriate ones. A longstanding question is how neuronal activity eliminates specific synapses based on their strength. The technical challenges of in vivo studies have made it difficult to identify a molecular link between decreased activity and synapse elimination. We developed an organotypic coculture model of the mouse retinogeniculate system that facilitates real-time imaging and elucidation of molecular mechanisms underlying the removal of less active synapses during synaptic competition. Using this model we show for the first time that complement component C1q is necessary for activity-dependent synaptic competition and preferentially localizes to less active, competing presynaptic inputs. In conjunction with classic in vivo and ex vivo models, this coculture model is a new tool to reveal molecular pathways underlying CNS circuit refinement.

scholarly journals Extra-gastric expression of the proton pump H+/K+-ATPase in the gills and kidney of the teleost Oreochromis niloticus

2020 ◽  
Vol 223 (16) ◽  
pp. jeb214890
Author(s):  
Ebtesam Ali Barnawi ◽  
Justine E. Doherty ◽  
Patrícia Gomes Ferreira ◽  
Jonathan M. Wilson
Keyword(s):  
Oreochromis Niloticus ◽  
Cellular Localization ◽  
Ex Vivo ◽  
Uptake Inhibition ◽  
Custom Made ◽  
Potassium Regulation ◽  
Immunohistochemical Techniques ◽  
Collecting Tubules ◽  
First Time

ABSTRACTPotassium regulation is essential for the proper functioning of excitable tissues in vertebrates. The H+/K+-ATPase (HKA), which is composed of the HKα1 (gene: atp4a) and HKβ (gene: atp4b) subunits, has an established role in potassium and acid–base regulation in mammals and is well known for its role in gastric acidification. However, the role of HKA in extra-gastric organs such as the gill and kidney is less clear, especially in fishes. In the present study in Nile tilapia, Oreochromis niloticus, uptake of the K+ surrogate flux marker rubidium (Rb+) was demonstrated in vivo; however, this uptake was not inhibited with omeprazole, a potent inhibitor of the gastric HKA. This contrasts with gill and kidney ex vivo preparations, where tissue Rb+ uptake was significantly inhibited by omeprazole and SCH28080, another gastric HKA inhibitor. The cellular localization of this pump in both the gill and kidney was demonstrated using immunohistochemical techniques with custom-made antibodies specific for Atp4a and Atp4b. Antibodies against the two subunits showed the same apical ionocyte distribution pattern in the gill and collecting tubules/ducts in the kidney. Atp4a antibody specificity was confirmed by western blotting. RT-PCT was used to confirm the expression of both subunits in the gill and kidney. Taken together, these results indicate for the first time K+ (Rb+) uptake in O. niloticus and that HKA is implicated, as shown through the ex vivo uptake inhibition by omeprazole and SCH28080, verifying a role for HKA in K+ absorption in the gill's ionocytes and collecting tubule/duct segments of the kidney.

Abstract P283: Parkin Mediates Mitophagy in Cardioprotection

2011 ◽  
Vol 109 (suppl_1) ◽  
Author(s):  
Allen M Andres ◽  
Chengqun Huang ◽  
Eric P Ratliff ◽  
Genaro Hernandez ◽  
Pamela Lee ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Wild Type ◽  
Simulated Ischemia ◽  
Selective Targeting ◽  
Cardioprotective Effects ◽  
Mitochondrial Turnover ◽  
First Time

Autophagy-dependent mitochondrial turnover in response to cellular stress is necessary for maintaining cellular homeostasis. However, the mechanisms that govern the selective targeting of damaged mitochondria are poorly understood. Parkin, an E3 ubiquitin ligase, has been shown to be essential for the selective clearance of damaged mitochondria. Parkin is expressed in the heart, yet its function has not been investigated in the context of cardioprotection. We previously reported that autophagy is required for cardioprotection by ischemic preconditioning (IPC). In the present study, we used simulated ischemia in vitro and IPC in hearts (in vivo and ex vivo) to investigate the role of Parkin in mediating cardioprotection. In HL-1 cells, simulated ischemia induced Parkin translocation to mitochondria and mitochondrial elimination. Mitochondrial loss was blunted in Atg5-deficient cells, revealing the requirement for autophagy in mitochondrial elimination. Consistent with previous reports implicating p62/SQSTM1 in mitophagy, we found that downregulation of p62 attenuated mitophagy and exacerbated cell death in HL-1 cardiomyocytes subjected to simulated ischemia. While wild type mice showed p62 translocation to mitochondria after IPC, Parkin knockout mice exhibited attenuated translocation of p62 to mitochondria. Importantly, ablation of Parkin in mice abolished the cardioprotective effects of IPC. These results reveal for the first time the crucial role of Parkin and mitophagy in cardioprotection.

scholarly journals Fishing for Targets of Alien Metabolites: A Novel Peroxisome Proliferator-Activated Receptor (PPAR) Agonist from a Marine Pest

Marine Drugs ◽  
2018 ◽  
Vol 16 (11) ◽  
pp. 431 ◽  
Author(s):  
Rosa Vitale ◽  
Enrico D'Aniello ◽  
Stefania Gorbi ◽  
Andrea Martella ◽  
Cristoforo Silvestri ◽  
...  
Keyword(s):  
Native Species ◽  
Molecular Mechanisms ◽  
Ex Vivo ◽  
Invasion Biology ◽  
In Vitro Assays ◽  
Bioactive Metabolites ◽  
Peroxisome Proliferator ◽  
Invasive Pests

Although the chemical warfare between invasive and native species has become a central problem in invasion biology, the molecular mechanisms by which bioactive metabolites from invasive pests influence local communities remain poorly characterized. This study demonstrates that the alkaloid caulerpin (CAU)—a bioactive component of the green alga Caulerpa cylindracea that has invaded the entire Mediterranean basin—is an agonist of peroxisome proliferator-activated receptors (PPARs). Our interdisciplinary study started with the in silico prediction of the ligand-protein interaction, which was then validated by in vivo, ex vivo and in vitro assays. On the basis of these results, we candidate CAU as a causal factor of the metabolic and behavioural disorders observed in Diplodus sargus, a native edible fish of high ecological and commercial relevance, feeding on C. cylindracea. Moreover, given the considerable interest in PPAR activators for the treatment of relevant human diseases, our findings are also discussed in terms of a possible nutraceutical/pharmacological valorisation of the invasive algal biomasses, supporting an innovative strategy for conserving biodiversity as an alternative to unrealistic campaigns for the eradication of invasive pests.

scholarly journals Microbial enzymes induce colitis by reactivating triclosan in the mouse gastrointestinal tract

2022 ◽  
Vol 13 (1) ◽  
Author(s):  
Jianan Zhang ◽  
Morgan E. Walker ◽  
Katherine Z. Sanidad ◽  
Hongna Zhang ◽  
Yanshan Liang ◽  
...  
Keyword(s):  
Gut Microbiota ◽  
Molecular Mechanisms ◽  
Ex Vivo ◽  
Metabolic Activation ◽  
Consumer Products ◽  
Environmental Chemicals ◽  
Intestinal Microbes ◽  
Targeted Inhibition

AbstractEmerging research supports that triclosan (TCS), an antimicrobial agent found in thousands of consumer products, exacerbates colitis and colitis-associated colorectal tumorigenesis in animal models. While the intestinal toxicities of TCS require the presence of gut microbiota, the molecular mechanisms involved have not been defined. Here we show that intestinal commensal microbes mediate metabolic activation of TCS in the colon and drive its gut toxicology. Using a range of in vitro, ex vivo, and in vivo approaches, we identify specific microbial β-glucuronidase (GUS) enzymes involved and pinpoint molecular motifs required to metabolically activate TCS in the gut. Finally, we show that targeted inhibition of bacterial GUS enzymes abolishes the colitis-promoting effects of TCS, supporting an essential role of specific microbial proteins in TCS toxicity. Together, our results define a mechanism by which intestinal microbes contribute to the metabolic activation and gut toxicity of TCS, and highlight the importance of considering the contributions of the gut microbiota in evaluating the toxic potential of environmental chemicals.

scholarly journals Transcriptomic analysis of equine chorioallantois reveals immune networks and molecular mechanisms involved in nocardioform placentitis

2021 ◽  
Vol 52 (1) ◽  
Author(s):  
Hossam El-Sheikh Ali ◽  
Shavahn C. Loux ◽  
Laura Kennedy ◽  
Kirsten E. Scoggin ◽  
Pouya Dini ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Glucose Transporter ◽  
Inflammasome Activation ◽  
Molecular Pathways ◽  
Toll Like Receptor ◽  
Inflammatory Signaling ◽  
Leukocytic Infiltration ◽  
Apoptosis Pathways ◽  
Placental Inflammation ◽  
First Time

AbstractNocardioform placentitis (NP) continues to result in episodic outbreaks of abortion and preterm birth in mares and remains a poorly understood disease. The objective of this study was to characterize the transcriptome of the chorioallantois (CA) of mares with NP. The CA were collected from mares with confirmed NP based upon histopathology, microbiological culture and PCR for Amycolatopsis spp. Samples were collected from the margin of the NP lesion (NPL, n = 4) and grossly normal region (NPN, n = 4). Additionally, CA samples were collected from normal postpartum mares (Control; CRL, n = 4). Transcriptome analysis identified 2892 differentially expressed genes (DEGs) in NPL vs. CRL and 2450 DEGs in NPL vs. NPN. Functional genomics analysis elucidated that inflammatory signaling, toll-like receptor signaling, inflammasome activation, chemotaxis, and apoptosis pathways are involved in NP. The increased leukocytic infiltration in NPL was associated with the upregulation of matrix metalloproteinase (MMP1, MMP3, and MMP8) and apoptosis-related genes, such as caspases (CASP3 and CASP7), which could explain placental separation associated with NP. Also, NP was associated with downregulation of several placenta-regulatory genes (ABCG2, GCM1, EPAS1, and NR3C1), angiogenesis-related genes (VEGFA, FLT1, KDR, and ANGPT2), and glucose transporter coding genes (GLUT1, GLUT10, and GLUT12), as well as upregulation of hypoxia-related genes (HIF1A and EGLN3), which could elucidate placental insufficiency accompanying NP. In conclusion, our findings revealed for the first time, the key regulators and mechanisms underlying placental inflammation, separation, and insufficiency during NP, which might lead to the development of efficacious therapies or diagnostic aids by targeting the key molecular pathways.

scholarly journals Molecular Mechanisms of Aroma Persistence: From Noncovalent Interactions Between Aroma Compounds and Oral Mucosa to Metabolization of Aroma Compounds by Saliva and Oral Cells

2021 ◽  
Author(s):  
Carolina Muñoz-Gonzalez ◽  
Marine Brulé ◽  
Christophe Martin ◽  
Gilles Feron ◽  
Francis Canon
Keyword(s):  
Oral Mucosa ◽  
Molecular Mechanisms ◽  
Ex Vivo ◽  
Noncovalent Interactions ◽  
Aroma Compounds ◽  
Integral Approach ◽  
Exhaled Air ◽  
Oral Cells ◽  
Novel Model

<p>Aroma persistence plays a major role in the liking and wanting of orally consumed products (food, dental toiletries, tobacco, drugs, etc.). Here, we use an integral approach including <i>ex vivo</i> experiments using a novel model of oral mucosa and saliva in well controlled conditions as well as <i>in vivo</i> dynamic instrumental and sensory experiments. <i>Ex vivo</i> experiments show the ability of the mucosal pellicle, the thin layer of salivary proteins covering the oral mucosa, to interact with aroma compounds, as well as the ability of oral cells and saliva to metabolize carbonyl aroma compounds. <i>In vivo</i> evaluation of the exhaled air and perception of individuals after aroma sample consumption confirm <i>ex vivo</i> findings in a more real context. Thus, aroma compounds susceptible to be metabolized by saliva and oral cells show a lower aroma persistence than non metabolized compounds, for which other mechanisms such as the adsorption at the surface of the oral mucosa (mucosal pellicle) as a function of their hydrophobicity are involved. Thus, we argue that the physiological aspects occurring during the oral processing, and especially, metabolization of aroma compounds, have to be considered when studying the phenomenon of aroma persistence.</p>

The expression, lamin-dependent localization and RNAi depletion phenotype for emerin inC. elegans

2002 ◽  
Vol 115 (5) ◽  
pp. 923-929 ◽  
Author(s):  
Yosef Gruenbaum ◽  
Kenneth K. Lee ◽  
Jun Liu ◽  
Merav Cohen ◽  
Katherine L. Wilson
Keyword(s):  
Nuclear Envelope ◽  
Molecular Mechanisms ◽  
Cell Types ◽  
Growth Conditions ◽  
Rnai Phenotype ◽  
Lamin B ◽  
C Elegans ◽  
Nuclear Lamins ◽  
First Time

Emerin belongs to the LEM-domain family of nuclear membrane proteins, which are conserved in metazoans from C. elegans to humans. Loss of emerin in humans causes the X-linked form of Emery-Dreifuss muscular dystrophy(EDMD), but the disease mechanism is not understood. We have begun to address the function of emerin in C. elegans, a genetically tractable nematode. The emerin gene (emr-1) is conserved in C. elegans. We detect Ce-emerin protein in the nuclear envelopes of all cell types except sperm, and find that Ce-emerin co-immunoprecipitates with Ce-lamin from embryo lysates. We show for the first time in any organism that nuclear lamins are essential for the nuclear envelope localization of emerin during early development. We further show that four other types of nuclear envelope proteins, including fellow LEM-domain protein Ce-MAN1, as well as Ce-lamin, UNC-84 and nucleoporins do not depend on Ce-emerin for their localization. This result suggests that emerin is not essential to organize or localize the only lamin (B-type) expressed in C. elegans. We also analyzed the RNAi phenotype resulting from the loss of emerin function in C. elegans under laboratory growth conditions, and found no detectable phenotype throughout development. We propose that C. elegans is an appropriate system in which to study the molecular mechanisms of emerin function in vivo.

scholarly journals Understanding the Effects of General Anesthetics on Cortical Network Activity Using Ex Vivo Preparations

2019 ◽  
Vol 130 (6) ◽  
pp. 1049-1063 ◽  
Author(s):  
Logan J. Voss ◽  
Paul S. García ◽  
Harald Hentschke ◽  
Matthew I. Banks
Keyword(s):  
Molecular Mechanisms ◽  
Ex Vivo ◽  
Brain Slices ◽  
Brain Regions ◽  
Cortical Network ◽  
Network Activity ◽  
Common Mechanism ◽  
General Anesthetics ◽  
Neural Basis

Abstract General anesthetics have been used to ablate consciousness during surgery for more than 150 yr. Despite significant advances in our understanding of their molecular-level pharmacologic effects, comparatively little is known about how anesthetics alter brain dynamics to cause unconsciousness. Consequently, while anesthesia practice is now routine and safe, there are many vagaries that remain unexplained. In this paper, the authors review the evidence that cortical network activity is particularly sensitive to general anesthetics, and suggest that disruption to communication in, and/or among, cortical brain regions is a common mechanism of anesthesia that ultimately produces loss of consciousness. The authors review data from acute brain slices and organotypic cultures showing that anesthetics with differing molecular mechanisms of action share in common the ability to impair neurophysiologic communication. While many questions remain, together, ex vivo and in vivo investigations suggest that a unified understanding of both clinical anesthesia and the neural basis of consciousness is attainable.

Pathologic up-regulation of TNFSF15–TNFRSF25 axis sustains endothelial dysfunction in unprovoked venous thromboembolism

2019 ◽  
Vol 116 (3) ◽  
pp. 698-707
Author(s):  
Silvia Della Bella ◽  
Francesca Calcaterra ◽  
Monica Bacci ◽  
Claudia Carenza ◽  
Chiara Pandolfo ◽  
...  
Keyword(s):  
Venous Thromboembolism ◽  
Molecular Mechanisms ◽  
Ex Vivo ◽  
Death Receptor ◽  
Lower Growth ◽  
Endothelial Colony Forming Cells ◽  
Healthy Donors ◽  
Novel Therapeutic Strategies ◽  
Microarray Expression

Abstract Aims The pathogenetic mechanisms underlying unprovoked venous thromboembolism (uVTE) are largely unknown. In this study, we investigated the molecular mechanisms involved in uVTE pathogenesis by using ex vivo expanded endothelial colony-forming cells (ECFCs), which represent a valuable non-invasive tool for the assessment of endothelial function. Methods and results We isolated and expanded ECFCs from the peripheral blood of uVTE patients and observed that these cells underwent earlier senescence and showed lower growth rate compared with ECFCs obtained from healthy donors. Through microarray expression profiling, we demonstrated that 2905 genes were differentially expressed between patients and controls. Among them, the anti-angiogenic cytokine TNF superfamily member 15 (TNFSF15) and its death-receptor TNFRSF25 were up-regulated in uVTE ECFCs, and this finding was validated by RT-qPCR. TNFSF15 up-regulation was confirmed at the protein level in ECFC supernatants, and the in vivo relevance of these findings was further corroborated by demonstrating that also the plasmatic levels of TNFSF15 are increased in uVTE patients. After proving that exogenous TNFSF15 exerts pro-apoptotic and anti-proliferative activity on control ECFCs, we demonstrated through blocking experiments that TNFSF15 up-regulation contributes to impaired survival and proliferation of uVTE ECFCs. Conclusion By providing evidence that TNFSF15 impairs ECFC functions crucial to endothelial repair, and that uVTE patients have increased TNFSF15 levels both ex vivo and in vivo, the results of this study suggest that pathologic up-regulation of TNFSF15–TNFRSF25 axis may contribute to uVTE pathogenesis, and may represent the target for novel therapeutic strategies aimed at preventing recurrences in uVTE patients.

scholarly journals Candida albicans forms biofilms on the vaginal mucosa

Microbiology ◽  
2010 ◽  
Vol 156 (12) ◽  
pp. 3635-3644 ◽  
Author(s):  
M. M. Harriott ◽  
E. A. Lilly ◽  
T. E. Rodriguez ◽  
P. L. Fidel ◽  
M. C. Noverr
Keyword(s):  
Biofilm Formation ◽  
Ex Vivo ◽  
Microscopic Analysis ◽  
Vaginal Mucosa ◽  
First Time ◽  
Established Role ◽  
Type Strains

Current understanding of resistance and susceptibility to vulvovaginal candidiasis challenges existing paradigms of host defence against fungal infection. While abiotic biofilm formation has a clearly established role during systemic Candida infections, it is not known whether C. albicans forms biofilms on the vaginal mucosa and the possible role of biofilms in disease. In vivo and ex vivo murine vaginitis models were employed to examine biofilm formation by scanning electron and confocal microscopy. C. albicans strains included 3153A (lab strain), DAY185 (parental control strain), and mutants defective in morphogenesis and/or biofilm formation in vitro (efg1/efg1 and bcr1/bcr1). Both 3153A and DAY815 formed biofilms on the vaginal mucosa in vivo and ex vivo as indicated by high fungal burden and microscopic analysis demonstrating typical biofilm architecture and presence of extracellular matrix (ECM) co-localized with the presence of fungi. In contrast, efg1/efg1 and bcr1/bcr1 mutant strains exhibited weak or no biofilm formation/ECM production in both models compared to wild-type strains and complemented mutants despite comparable colonization levels. These data show for the first time that C. albicans forms biofilms in vivo on vaginal epithelium, and that in vivo biotic biofilm formation requires regulators of biofilm formation (BCR1) and morphogenesis (EFG1).

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